ABSTRACT Acute lymphoblastic leukemia (ALL), the most common malignancy of childhood, has a suspected prenatal origin in a majority of cases. Although nearly 90% of ALL patients survive into adulthood, treatment has devastating long-term health effects and primary prevention remains the quintessential goal of oncology research. Children who develop ALL exhibit alterations in inflammatory cytokine levels at birth and experience more medically diagnosed early-life infections, suggesting that early-life infections may be a modifiable etiologic agent. We recently demonstrated that pretreatment bone marrow specimens from children with ALL had prevalent cytomegalovirus (CMV) infection. Screening of archived newborn blood samples from 268 children who went on to develop ALL and 270 cancer-free control children demonstrated that ALL patients were nearly 4-times more likely to have detectable CMV in their blood at birth (OR=3.71, P=0.0016), suggesting that congenital CMV infection is an ALL risk factor. CMV is the most common congenital infection worldwide, affecting 1 in 150 infants. CMV infection prior to birth likely has a significant impact on the developing immune system, including T cell activation and central tolerance. CMV has the largest genome of any human viral pathogen and harbors many immune-evasion genes, indicating that CMV modulates the host immune system to escape immunosurveillance. The mechanisms underlying this CMV-induced immune modulation are poorly understood; however, other oncogenic viruses can induce widespread methylation changes to the host epigenome. Virally-induced epigenetic changes may alter the transcriptional landscape of the developing immune system and negatively impact both lineage commitment during hematopoiesis and host immunosurveillance, thereby augmenting ALL risk. Thus, defining the interaction between congenital CMV infection and dysregulation of leukemia-associated genes in early life is an important step toward establishing the mechanistic link between congenital CMV infection and ALL risk, a potentially vaccine-preventable cancer risk factor. We hypothesize that congenital CMV infection induces epigenetic and immunologic changes in the developing fetus that contribute to risk of developing ALL during childhood. Using a matched case-control sample of CMV-infected, CMV-exposed, and CMV-unexposed cord blood donors from the Carolinas Cord Blood Bank, we will identify the epigenetic and immunologic consequences of congenital CMV infection. We will then determine if CMV infection-associated epigenetic and immunologic changes are recapitulated in newborn blood spots from children who went on to develop ALL. Finally, we will determine whether known ALL susceptibility variants modify risk of congenital CMV infection by comparing the frequency of these variants in CMV-infected cord blood donors to uninfected controls. These studies will elucidate the role of congenital CMV infection on later cancer risk and help to identify modifiable early-life factors that can reduce the public health burden associated with the most common cancer of childhood.